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The protein kinase Akt plays a pivotal role in cellular processes. However, its isoforms' distinct functions have not been resolved to date, mainly due to the lack of suitable biochemical and cellular tools. Against this background, we present the development of an isoform-dependent Ba/F3 model system to translate biochemical results on isoform specificity to the cellular level. Our cellular model system complemented by protein X-ray crystallography and structure-based ligand design results in covalent-allosteric Akt inhibitors with unique selectivity profiles. In a first proof-of-concept, the developed molecules allow studies on isoform-selective effects of Akt inhibition in cancer cells. Thus, this study will pave the way to resolve isoform-selective roles in health and disease and foster the development of next-generation therapeutics with superior on-target properties., (© 2021. The Author(s).)

The identification of compounds for dissecting biological functions and the development of novel drug molecules are central tasks that often require screening campaigns. However, the required architecture is cost- and time-intensive. Herein we describe the devices and technologies that comprise a Robotics-Assisted Screening Platform for Efficient Ligand Discovery (RASPELD), which we set up in an academic laboratory. RASPELD provides semi-automated high-end screening, and it can be maintained by graduate students. We demonstrate its successful application in biochemical and cellular screens for the identification and validation of bioactive chemical entities as candidate cancer-relevant inhibitors. Specifically, we examined the interaction between a transcription factor, Nrf2, and its key regulator, Keap1. We also examined drug-resistant mutants of the epidermal growth factor receptor (EGFR). Screening campaigns with more than 30 000 compounds were performed in a reasonable period of time. We identified the molecule RSL6586 as a starting point for hit optimization, which is currently ongoing., (© 2018 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.)